How to Build Plasmid Engineering Workbench with Circular Mapping, Restriction Analysis, Virtual Gels, and Primer Design

In this tutorial, we build a Google Colab-native plasmid workbench that recreates the core ideas of SpliceCraft inside an interactive notebook environment. Instead of relying on a terminal-based TUI, we use Biopython, NumPy, and Matplotlib to load plasmid records, normalize annotated genomic features, render circular and linear plasmid maps, compute sequence statistics, analyze restriction enzyme cut sites, simulate virtual digests, scan open reading frames, translate CDS features, design primers, and apply sequence edits programmatically. We begin with a synthetic offline plasmid so the workflow runs reliably without external dependencies, while still supporting optional NCBI GenBank fetching and local GenBank uploads.

We set up the notebook environment, install Biopython when needed, and import the scientific, plotting, and sequence-analysis libraries required for the workflow. We define the plasmid configuration, feature color palette, multiple cloning site, and fallback synthetic plasmid so the tutorial works even without an internet connection. We also create helper functions to load records from NCBI or GenBank files and normalize annotated sequence features into drawable metadata.

We implement the circular plasmid visualization by mapping base-pair positions to angular coordinates on a clockwise ring. We calculate GC content, draw the plasmid backbone, add tick marks, render feature arcs, and attach directional arrowheads to clearly indicate strand orientation. We also add labels and central sequence metadata so that the map is both biologically informative and visually readable in Colab.

We build a linear plasmid map that represents the same annotated features along a base-pair axis for easier positional inspection. We then compute the cumulative GC-skew to observe directional nucleotide bias that can indicate replication-related structure. We also add sequence statistics and restriction-analysis utilities to summarize plasmid composition, enzyme cut sites, unique cutters, and expected digest fragments.

We simulate a virtual agarose gel by plotting digest fragment sizes beside a DNA ladder on a log-scaled migration axis. We scan the plasmid sequence in all six reading frames to identify long open reading frames and rank them by amino-acid length. We also translate annotated CDS features and design primers around a selected region by tuning primer length toward a target melting temperature.

We implement a pure sequence-editing function that supports insertion, deletion, and replacement while returning a new edited SeqRecord. We preserve plasmid annotations and shift downstream feature coordinates so the edited construct remains internally consistent after sequence changes. We also create a lightweight in-notebook plasmid library that lets us store and list original and edited records during the workflow.